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Yeast biodiversity in the guts of several pests on maize; comparison of three methods: classical isolation, cloning and DGGE

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Abstract

The yeast biodiversity in the guts of several pests (Diabrotica virgifera, Helicoverpa armigera, Ostrinia nubilalis) on maize from two isolation sources was assessed by cultivation-dependent and cultivation-independent methods. These yeasts are considered to bear a potentially high biotechnological relevance due to their potential ability to degrade several mycotoxins incorporated by their hosts. The 97 isolated yeast strains showed 21 different partial sequence types of the 26S rRNA gene which could be assigned to 10 different genera. The determined genera and species are discussed in terms of the meaning of their taxonomic status or their occurrence in nature. Two cultivation-independent methods, cloning and DGGE, were compared. We propose the combination of these methods as well as the combination of both cultivation-independent and cultivation-dependent approaches, for gaining better insights into fungal biodiversity.

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Abbreviations

DGGE:

denaturing gradient gel electrophoresis

TGGE:

temperature gradient gel electrophoresis

SSCP:

single strand conformation polymorphism

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Acknowledgements

We are grateful to Assistant Prof. Joseph Strauss and Dr. Markus Gorfer for accomplishing the cloning experiments in their laboratory. We thank to Dr. Peter Cate and Helmut Klapal who helped us to collect and identify the insects in the fields.

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Authors and Affiliations

  1. Austrian Center of Biological Resources and Applied Mycology (ACBR), Institute of Applied Microbiology, University of Natural Resources and Applied Life Sciences, Muthgasse 18, 1190, Vienna, Austria

    Orsolya Molnár, Michael Wuczkowski & Hansjörg Prillinger

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  1. Orsolya Molnár
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  2. Michael Wuczkowski
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  3. Hansjörg Prillinger
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Correspondence to Hansjörg Prillinger.

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Fig. S1

Hanseniaspora uvarum, budding cells, two days on GYP agar. Bar indicates 10 μm. (GIF 36.4 kb)

Fig. S2

Tilletiopsis washingtonensis. A 2 days on cornmeal agar, note the sterigmata (arrows); B ballistoconidia. Bars indicate 10 μm. (GIF 44.9 kb)

Fig. S3

PCR-fingerprints of strains identified as Pichia guilliermondii on the basis of their rDNA sequences. Amplification primed by M13 (A), (GTG)5 (B) and V5 (C). M = length marker. CBS 2030T is the type strain of Pichia guilliermondii, CBS 5975T is the type strain of Candida xestobii and CBS 7921T is the type strain of Candida fukuyamaensis. Note the similarities of the strains identified as P. guilliermondii (HA 1658-HA 1663) and C. xestobii (HA 1664-HA 1668) amplified with all the three primers. (GIF 127 kb)

Fig. S4

PCR-fingerprints of strains identified as Aureobasidium pullulans on the basis of their rDNA sequences. Amplification primed by M13 (A), (GTG)5 (B, lanes 2–4) and V5 (B, lanes 5–7). M Length marker. CBS 584.75 is a confirmed strain of A. pullulans. (GIF 25.6 kb)

Fig. S5

PCR-fingerprints of strains identified as Candida quercitrusa. Amplification primed by M13 (lanes 2–3), (GTG)5 (lanes 4–5) and V5 (lanes 6–7). M Length marker. CBS 4412T is the type strain of C. quercitrusa. (GIF 28.4 kb)

Fig. S6

PCR-fingerprints of strains identified as Hanseniasporum uvarum. Amplification primed by M13 (lanes 1–2), (GTG)5 (lanes 3–4) and V5 (lanes 6–7). M   Length marker. CBS 314T is the type strain of H. uvarum. (GIF 29.0 kb)

Fig. S7

PCR-fingerprints of strains identified as Rhodotorula glutinis. Amplification primed by M13 (lanes 1–6) and V5 (lanes 8–13). M Length marker. CBS 20T is the type strain of R. glutinis. CBS 6085T is the type strain of Rhodosporidium diobovatum. (GIF 22.0 kb)

Fig. S8

PCR-fingerprints of strains identified as Sporobolomyces coprosmae. Amplification primed by M13 (lanes 1–4) and V5 (lanes 6–9). M Length marker. CBS 7899T is the type strain of S. coprosmae. (GIF 31.9 kb)

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Molnár, O., Wuczkowski, M. & Prillinger, H. Yeast biodiversity in the guts of several pests on maize; comparison of three methods: classical isolation, cloning and DGGE. Mycol Progress 7, 111–123 (2008). https://doi.org/10.1007/s11557-008-0558-0

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